Conventionally, in HPLC (high performance liquid chromatography) analysis, isocratic analysis in which elution occurs with a solvent of a fixed composition and gradient analysis in which elution occurs while changing with time a mobile phase composition are employed. In the isocratic analysis, a stable chromatogram is obtained. However, in the isocratic analysis, it sometimes takes a long time to resolve components which are different in the retentivity to a stationary phase, or the peak may be broadened. In contrast, in the gradient analysis, by changing the composition ratio of a mobile phase and increasing an elution ability, analysis time can be shortened and broadening of the peak can be suppressed.
Various resolving agents and columns are commercially available as a stationary phase for resolving optical isomers. As such resolving agents for optical isomers, used frequently are polysaccharide derivatives which demonstrates excellent resolution properties in resolving various compounds. Moreover, in recent years, a solvent-resistant resolving agent for optical isomers has been developed which can be used under mobile phase conditions of a polar solvent and, in which isomers a polysaccharide derivative has been immobilized on a carrier (Patent Documents 1 and 2). A column for resolving optical isomers, which has been charged with the polysaccharide-based solvent-resistant resolving agent, has drawn attention because various solvents has become available as a mobile phase while high resolution properties of the polysaccharide derivatives are maintained. However, a trial-and-error procedure is required for achieving sharp resolution in a short time (preferably 30 minutes or shorter).
Patent Document 1: JP 2751003 A
Patent Document 2: JP 2751004 A